BASIC HELIX-LOOP-HELIX TRANSCRIPTION FACTOR <EM>TCF4</EM> IS A KEY REGULATOR OF STIMULUS-DEPENDENT GENE EXPRESSION IN NEURONS

Transcription Factor 4 (TCF4) is a basic helix–loop–helix transcription factor that regulates neuronal differentiation and signalling. TCF4 has been implicated in schizophrenia and other neuropsychiatric disorders and is the causative gene for the autism spectrum disorder Pitt–Hopkins syndrome. TCF4 functions through dimerization with other transcription factors, and its transcriptional activity is regulated by neuronal activity via cAMP signalling. However, the broader role of TCF4 in activity-dependent transcriptional regulation and its regulatory network in primary neurons remains poorly understood. To address this, we mapped genome-wide TCF4 binding under depolarizing conditions using chromatin immunoprecipitation sequencing with two antibodies recognizing all TCF4 isoforms. We identified three predominant TCF4-binding E-box motifs in rat primary cortical neurons, with binding preferences dependent on TCF4 interaction partners. To uncover regulators of the E-box–dependent regulatory landscape in response to depolarization, we performed in vitro DNA pulldown assays using these motifs, enabling identification and validation of novel TCF4 interactors and co-regulators. Finally, RNA sequencing following Tcf4 knockdown in depolarized primary neurons revealed alterations in activity-dependent gene expression programs. Collectively, our findings provide insight into the stimulus-dependent TCF4 regulatory network and its role in neuronal homeostasis and neurodevelopmental disorders.